The present invention relates to the field of chemistry and more specifically to the field of peptide chemistry. More precisely, it is directed to acyl-dipeptide-like compounds bearing an accessory functionally substituted side chain spacer which can optionally be grafted in the form of conjugates, the accessory side chain spacer of which further imparts original properties to the molecule in terms of biological activity and physical chemical characteristics. Depending on the chemical species involved, the accessory side chain spacer gives the acyl-dipeptide-like compound added functional ability by finely tuning its original properties and by conferring novel ones thereto as well. These molecules bearing an accessory functional side chain spacer can be conjugated to a pharmaceutical carrier, an antigen or a vehicle. Bioconjugation involves coupling two or more chemical species to form a novel molecular complex having properties differing from those of the individual components. Natural or synthetic products, having inherent pharmacological properties, can be mutually combined to make new species having original or improved pharmacological and chemical physical properties as compared to starting compounds. Bioconjugates have a wide range of applications in all fields of human medicine and animal care as well as diagnostics.
A great number of homo- or heterobifunctional coupling agents have already been described and may be used in coupling molecules ranging from amino acids, to peptides, protein, sugars, oligosaccharides, polysaccharides, nucleic acids, oligonucleotides, polynucleotides, lipids, and nearly every single molecule bearing a functional group capable of bonding. Considerable effort has been made in recent years regarding the synthesis of antigenic constructs made from two molecules bearing different messages. Good et al. [(1987), Science, 235: 1059-1062], have for instance reported the synthesis of a peptide containing both T helper and B lymphocyte recognition epitopes. Bessler and Jung [(1992) Res. Immunol., 5: 548-553] have disclosed conjugates composed of a peptide and an immunostimulant. Hoffmann et al. [(1997) FEMS Immunol. Med. Microbiol., 17: 225-234] have disclosed conjugates of a lipopeptide and a mellitin derived-synthetic peptide. Ulrich and Meyers [(1995) Vaccine Design, Plenum Press, New York, 495-524], have observed that immune response was inefficient unless the hapten and MPL adjuvant (Monophosphoryl Lipid A) were found in the same liposome. They suggested the possible existence of a covalent bond between MPL and the hapten. In fact, a hapten-adjuvant conjugate may prove to be highly efficient when used as a vaccine adjuvant. Ikeda et al. [(1999) Chem. Pharm. Bull., 47 (4), 563-568] have reported synthesis of a structural analog of Lipid A coupled to a peptide tumour-derived antigen and demonstrated it has in vitro mitogenic activity.
The conjugation concept may equally apply to protein or even protein polysaccharide conjugates. Indeed, it is well known that use of polysaccharides alone as a vaccine does only give rise to a weak immune response in children aged below 5, as no T cell-mediated response is involved. [Gotschlich et al. (1977); Antibodies in Human Diagnosis and Therapy, Peltola et al., (1977), Pediatrics 60: 730-737]. By contrast, linking polysaccharides to protein carriers does result in a much stronger immune response. This phenomenon was discovered in 1931 by Avery and Goebel [(1931), J. Exp. Med., 54: 437-447]. A variety of recently developed vaccines reflect progress accomplished so far in this field. Mention should be made of vaccines to Haemophilus influenzae and different serotypes of Streptococcus pneumoniae [Powell and Newman, (1995), Vaccine Design, Plenum Press, New York]. In the latter case, a multivalent vaccine has been developed [Sood et Fatton, (1998), Exp. Opin. Invest. Drugs, 7: (3), 333-347].
A complex composed of an adjuvant bound to a polysaccharide-protein unit raises new opportunities. Chemical synthesis technology as applied to bioconjugates is currently well developed and one can carry out a number of projects which were inconceivable just a few years ago, relying on the wide range of homo- or heterobifunctional reagents now available and using conjugation procedures extensively reported in the literature [Hermanson, (1996), Bioconjugate Techniques, Academic Press, New York].
Thus, mention can be made, for instance, of the reductive amination method [Roy et al., (1984), Canad. J. Biochem. Cell Biol., 62: 270-279, Hermansson, p. 472] allowing conjugation of a carrier molecule to an aldehyde functional group, having a primary amine on a peptide or protein molecule, with a protein-polysaccharide conjugate or with a pharmaceutical carrier. This reaction results in the formation of a very stable dialkylamine compound.